rs758493597

Positions:

• chr19-11128089-A-G

Variant summary

Our verdict is Uncertain significance. Variant got 4 ACMG points: 4P and 0B. PM2 PP3 PP4

This summary comes from the ClinGen Evidence Repository: NM_000527.5(LDLR):c.2389+4A>G variant is classified as variant of Uncertain significance for Familial Hypercholesterolemia by applying evidence codes (PM2, PP3 and PP4) as defined by the ClinGen Familial Hypercholesterolemia Expert Panel LDLR-specific variant curation guidelines (https://doi.org/10.1101/2021.03.17.21252755). The supporting evidence is as follows: PM2 - PopMax MAF = 0.000008798 (0.0009%) in European non-Finnish (gnomAD v2.1.1). PP3 - No REVEL, splicing evaluation required. Functional data on splicing not available. A) variant located at -3 to +6 from canonical donor splice site Ratio variant/wt canonical acceptor/donor = 0,786004057 ---- It is below 0.8PP4 - Variant meets PM2. Identified in 1 FH case from Ambry Genetics who fulfills Simon-Broome criteria. LINK:https://erepo.genome.network/evrepo/ui/classification/CA040267/MONDO:0007750/013

• Frequency: Genomes: not found (cov: 31)
• Consequence: LDLR ENST00000558518.6 splice_donor_region, intron
• Scores: Splicing: ADA: 0.6781
• Clinical Significance: Uncertain significance reviewed by expert panel P:2 U:4 B:2
• Conservation: PhyloP100: 4.56

Genes affected

LDLR (HGNC:6547): (low density lipoprotein receptor) The low density lipoprotein receptor (LDLR) gene family consists of cell surface proteins involved in receptor-mediated endocytosis of specific ligands. The encoded protein is normally bound at the cell membrane, where it binds low density lipoprotein/cholesterol and is taken into the cell. Lysosomes release the cholesterol, which is made available for repression of microsomal enzyme 3-hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase, the rate-limiting step in cholesterol synthesis. At the same time, a reciprocal stimulation of cholesterol ester synthesis takes place. Mutations in this gene cause the autosomal dominant disorder, familial hypercholesterolemia. Alternate splicing results in multiple transcript variants.[provided by RefSeq, May 2022]

ACMG classification

Verdict is Uncertain_significance. Variant got 4 ACMG points.

• PM2: For more information check the summary or visit ClinGen Evidence Repository.
• PP3: For more information check the summary or visit ClinGen Evidence Repository.
• PP4: For more information check the summary or visit ClinGen Evidence Repository.

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	MANE	Protein	UniProt
LDLR	NM_000527.5	c.2389+4A>G		splice_donor_region_variant, intron_variant		ENST00000558518.6	NP_000518.1

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Protein	Appris	UniProt
LDLR	ENST00000558518.6	c.2389+4A>G		splice_donor_region_variant, intron_variant		1	NM_000527.5	ENSP00000454071	P3

Frequencies

GnomAD3 genomes Cov.: 31

GnomAD3 exomes AF: 0.00000398 AC: 1 AN: 251356 Hom.: 0 AF XY: 0.00 AC XY: 0 AN XY: 135888

Gnomad AFR exome AF: 0.00

Gnomad AMR exome AF: 0.00

Gnomad ASJ exome AF: 0.00

Gnomad EAS exome AF: 0.00

Gnomad SAS exome AF: 0.00

Gnomad FIN exome AF: 0.00

Gnomad NFE exome AF: 0.00000880

Gnomad OTH exome AF: 0.00

GnomAD4 exome Cov.: 32

GnomAD4 genome Cov.: 31

ClinVar

Significance: Uncertain significance

Submissions summary: Pathogenic:2 Uncertain:4 Benign:2

Revision: reviewed by expert panel

Submissions by phenotype

Hypercholesterolemia, familial, 1 Uncertain:3 Benign:2

Uncertain significance, criteria provided, single submitter	research	Fundacion Hipercolesterolemia Familiar	Mar 01, 2016	- -
Likely benign, criteria provided, single submitter	literature only	LDLR-LOVD, British Heart Foundation	Mar 25, 2016	- -
Benign, no assertion criteria provided	research	Laboratorium voor Moleculaire Diagnostiek Experimentele Vasculaire Geneeskunde, Academisch Medisch Centrum	-	- -
Uncertain significance, reviewed by expert panel	curation	ClinGen Familial Hypercholesterolemia Variant Curation Expert Panel	Jun 22, 2021	NM_000527.5(LDLR):c.2389+4A>G variant is classified as variant of Uncertain significance for Familial Hypercholesterolemia by applying evidence codes (PM2, PP3 and PP4) as defined by the ClinGen Familial Hypercholesterolemia Expert Panel LDLR-specific variant curation guidelines (https://doi.org/10.1101/2021.03.17.21252755). The supporting evidence is as follows: PM2 - PopMax MAF = 0.000008798 (0.0009%) in European non-Finnish (gnomAD v2.1.1). PP3 - No REVEL, splicing evaluation required. Functional data on splicing not available. A) variant located at -3 to +6 from canonical donor splice site Ratio variant/wt canonical acceptor/donor = 0,786004057 ---- It is below 0.8 PP4 - Variant meets PM2. Identified in 1 FH case from Ambry Genetics who fulfills Simon-Broome criteria. -
Uncertain significance, criteria provided, single submitter	research	Iberoamerican FH Network	Mar 01, 2016	- -

Familial hypercholesterolemia Pathogenic:1 Uncertain:1

Uncertain significance, criteria provided, single submitter	clinical testing	Labcorp Genetics (formerly Invitae), Labcorp	Oct 04, 2018	This sequence change falls in intron 16 of the LDLR gene. It does not directly change the encoded amino acid sequence of the LDLR protein, but it affects a nucleotide within the consensus splice site of the intron. This variant is present in population databases (rs758493597, ExAC 0.002%). This variant has been observed in several individuals affected with familial hypercholesterolemia (PMID: 15241806). ClinVar contains an entry for this variant (Variation ID: 252304). Nucleotide substitutions within the consensus splice site are a relatively common cause of aberrant splicing (PMID: 17576681, 9536098). Algorithms developed to predict the effect of sequence changes on RNA splicing suggest that this variant may disrupt the consensus splice site, but this prediction has not been confirmed by published transcriptional studies. In summary, the available evidence is currently insufficient to determine the role of this variant in disease. Therefore, it has been classified as a Variant of Uncertain Significance. -
Likely pathogenic, criteria provided, single submitter	clinical testing	Women's Health and Genetics/Laboratory Corporation of America, LabCorp	Nov 15, 2023	Variant summary: LDLR c.2389+4A>G alters a conserved nucleotide located close to a canonical splice site and therefore could affect mRNA splicing, leading to a significantly altered protein sequence. Several computational tools predict a significant impact on normal splicing: Four predict the variant weakens a 5' donor site. Four predict the variant creates a 3' acceptor site. Additionally, mRNA from carriers of this variant showed an additional band which PCR confirmed was lacking exon 16 (Aparicio_2023). The variant allele was found at a frequency of 4e-06 in 251356 control chromosomes. c.2389+4A>G has been reported in the literature in individuals affected with Familial Hypercholesterolemia in Spanish cohorts and registries, however these reports are lacking full clinical details of the individuals carrying the variant (Mozas_2004, Merino_2007, Alonso_2008, Martin-Campos_2018, Marco-Benedi_2022). However, a recent report showed that carriers of this variant have a high prevalence of familial history of premature ASCVD and hypercholesterolemia as well as significantly worse LDLc levels than the rest of the genetically confirmed FH cohort. In this cohort, a female carrier presented tendon xanthoma and a male carrier presented both corneal arcus and tendon xanthomas (Aparicio_2023). This last report provides strong evidence for causality. Six submitters have cited clinical-significance assessments for this variant to ClinVar after 2014. Multiple submitters reported the variant with conflicting assessments. Four labs, including the ClinGen Familial Hypercholesterolemia Variant Curation Expert Panel, classified the variant as VUS while one classified as likley benign and one classified as pathogenic. It is important to note, however, that all clinvar submitters have last updated their evidence prior to the recent report from Aparicio. Based on the evidence outlined above, the variant was classified as likely pathogenic. -

Cardiovascular phenotype Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

Ambry Genetics

Apr 09, 2021

The c.2389+4A>G pathogenic mutation results from an A to G substitution 4 nucleotides after coding exon 16 in the LDLR gene. This variant has been identified in several cohorts of individuals with familial hypercholesterolemia in Spain; however, clinical information is limited (Alonso R et al. Clin. Biochem., 2009 Jun;42:899-903; Mozas P et al. Hum. Mutat., 2004 Aug;24:187; Bourbon M et al. Atherosclerosis, 2017 07;262:8-13; Ibarretxe D et al. Atherosclerosis, 2018 11;278:210-216). This variant co-segregated with disease in one family tested in our laboratory, and RNA studies have demonstrated that this alteration results in abnormal splicing in the set of samples tested (Ambry internal data). The resulting transcript is in-frame and is not expected to trigger nonsense-mediated mRNA decay; however, the impacted region is critical for protein function (Ambry internal data). Other pathogenic alterations impacting the same donor site (c.2389G>T and c.2389+1G>T) have been shown to have a similar impact on splicing (Bourbon M et al. J. Med. Genet., 2009 May;46:352-7; Holla ØL et al. Mol. Genet. Metab., 2009 Apr;96:245-52). Based on the supporting evidence, this alteration is interpreted as a disease-causing mutation. -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
BayesDel_noAF	Benign	-0.50
CADD	Benign	18
DANN	Benign	0.78

Splicing

Name	Calibrated prediction	Score	Prediction
dbscSNV1_ADA	Benign	0.68
dbscSNV1_RF	Benign	0.54
SpliceAI score (max)		0.21		Details are displayed if max score is > 0.2
DS_DL_spliceai		0.21		Position offset: -4

Find out detailed SpliceAI scores and Pangolin per-transcript scores at spliceailookup.broadinstitute.org

Publications

LitVar

Other links and lift over

dbSNP: rs758493597; hg19: chr19-11238765;